Monoallelic gene expression, or allelic exclusion, is critical to the functioning of the immune system as it allows each lymphocyte to elaborate an antigen receptor of a single type. Chaos in immune system regulation might ensue if, for example, a B cell expressed both an antibody that responded to a pathogen and a second antibody that would cause damage to a certain host system. It has been proposed that the major pathway for implementing this choice is by a feedback mechanism in which the generation of a gene product from one allele leads to inhibition of the recombination machinery, thereby preventing rearrangement on the remaining germ line allele. While this mechanism may certainly play a maintenance role in inhibiting rearrangement on the non-selected allele, recent evidence suggests that the process of allelic exclusion may actually begin early in development, at about the time of implantation, when the antigen receptor genes become asynchronously replicating in each cell. However, it is not yet known how this chromosomal mark enables the initial selection of only one allele. We have designed experiments using targeted transgenic mice to show that replication timing plays a critical and direct role in controlling allelic exclusion. We will identify the molecular components that set up the asynchronous pattern of replication. We have developed a unique clonal pre-B cell system that allows us to decipher the monoallelic epigenetic processes such as chromatin modifications, unequal nuclear localization, DMA methylation, and replication that leads to allelic exclusion. These epigenetic mechanisms may be very similar to those that regulate the X-chromosome inactivation in female cells and parental genomic imprinting. These studies will help us to understand how the diversity and specificity of the immune system are generated and how this system functions to eliminate pathogens.